Listeria monocytogenes: the silent assassin

Graphical abstract Listeria monocytogenes is ubiquitous in both plant and animal reservoirs. It can persist in food production environments due to its capacity to grow at refrigerated temperatures and its resistance to biocides. The source of most human infections is contaminated food. Healthy individuals present with mild gastrointestinal symptoms. However, in immunocompromised individuals the infection is more severe, causing bacteraemia, meningitis and, in pregnancy-associated listeriosis, miscarriage and stillbirth. In vulnerable groups, including the elderly, pregnant women and their infants, listeriosis has a 20–30% mortality rate.


Graphical abstract
Listeria monocytogenes is ubiquitous in both plant and animal reservoirs.It can persist in food production environments due to its capacity to grow at refrigerated temperatures and its resistance to biocides.The source of most human infections is contaminated food.Healthy individuals present with mild gastrointestinal symptoms.However, in immunocompromised individuals the infection is more severe, causing bacteraemia, meningitis and, in pregnancy-associated listeriosis, miscarriage and stillbirth.In vulnerable groups, including the elderly, pregnant women and their infants, listeriosis has a 20-30% mortality rate.

MICROBIAL CHARACTERISTICS OF PHENOTYPIC/GENOTYPIC FEATURES
L. monocytogenes is a Gram-positive, non-spore-forming, motile, facultatively anaerobic, rod-shaped bacterium with peritrichous flagella.It is an intracellular pathogen capable of surviving in macrophages.Based on phylogenetic analysis, L. monocytogenes is divided into four major lineages (I-IV).Serotypes within Lineage I (1/2a, 3b, and 4b) are more commonly associated from human cases, while Lineage II serotypes (1/2c and 3a) are more often recovered from food and food production environments.Lineages III and IV are associated with animals [3,4].The ability to survive at low temperatures, high salt concentrations and low pH levels, and to form biofilms, facilitates survival and persistence in food and food processing environments [5].L. monocytogenes is ubiquitous in nature, in the animal reservoir and in plant, water, and soil environments.

CLINICAL DIAGNOSIS, LABORATORY CONFIRMATION AND SAFETY Clinical diagnosis
A clinical diagnosis is confirmed by the culture of L. monocytogenes from blood or cerebrospinal fluid, in patients presenting with sepsis or meningitis.Colonies are 0.5 to 1.5mm in diameter, smooth, translucent with a characteristic ground glass appearance able to be emulsified and with a zone of hazy β-haemolysis extending 1 to 2mm from the edge of the colony.Some strains of L. monocytogenes can be non-haemolytic.In pregnancy-associated listeriosis L. monocytogenes is cultured from vaginal swabs, placenta, meconium, or amniotic fluid.

Laboratory confirmation
Under the microscopy, L. monocytogenes appears as Gram positive rods approximately 0.5×0.5 -3 µm with rounded ends, occurring singly or sometimes in pairs and may resemble 'coryneforms' or diplococci.They are non-sporing, non-branching and non-capsulated.

Abstract
Listeriosis is a foodborne infection in humans caused by Listeria monocytogenes.Consumption of contaminated food can lead to severe infection in vulnerable patients, that can be fatal.Clinical manifestations include sepsis and meningitis, and in pregnancy-associated infection, miscarriage and stillbirth.Diagnosis is confirmed by culture and identification of the pathogen from blood, cerebrospinal fluid, vaginal swab, placenta or amniotic fluid.Treatment regimens recommend amoxicillin, ampicillin or an aminoglycoside.Virulence factors mediate bacterial adhesion and invasion of gut epithelial cells.Other factors mediate biofilm formation and tolerance to low temperatures and high salt concentrations facilitating persistence and survival in the environment.
Preliminary identification tests include a positive catalase reaction, tumbling motility at 20-25 °C, β-haemolysis and aesculin hydrolysis.Historically, the confirmatory test was the Christie-Atkins-Munch-Peterson (CAMP) test, which can help confirm species by testing for haemolysis enhancement on sheep blood agar.In recent years, identification may be confirmed by MALDI-ToF or PCR and/or whole genome sequencing [6].

Safety
L. monocytogenes is classified as a medium-risk biological organism and falls within Containment Level 2 (CL2) or Biosafety level 2 (BSL2) parameters.CL2/BSL2 laboratory protocols recommend that all bacterial manipulations be performed in a microbiological safety cabinet.Other interventions are focused on preventing injuries such as skin breakages, ingestion, and mucous membrane exposures.Pregnant women and immunocompromised staff are also strongly advised not to work with L. monocytogenes due to the potential for severe clinical outcomes in vulnerable groups.

TREATMENT, RESISTANCE AND EVASION Treatment
Treatment regimens recommend amoxicillin, ampicillin or an aminoglycoside [7].Listeriosis, manifesting as meningitis or septicaemia, requires intravenous amoxicillin/ampicillin.Trimethoprim-sulfamethoxazole (TMP-SMX) can be an alternative for penicillin allergy patients.For life-threatening infections, such as meningitis, gentamycin may be included.

Resistance
Although antimicrobial resistance in L. monocytogenes is low, resistance to penicillin, ampicillin, tetracycline, macrolides, and fluoroquinolones linked to efflux pumps been reported.Resistance to various disinfectants, such as benzalkonium chloride, is common and enables L. monocytogenes to persist in many food production environments [7].

Evasion
L. monocytogenes is an intracellular pathogen that can survive and replicate within host cells.The bacteria enter the host's cells through phagocytosis/endocytosis.The bacteria escapes phagosome using listeriolysin O to replicate in the cytoplasm.More specifically, L. monocytogenes can highjack the host cells enzymes and compounds for actin production, which helps them propel from cell to cell, evading the immune system [8].

PATHOGENIC STRATEGIES (HOST RANGE, HOST RESPONSE, TRANSMISSION, INFECTION AND VIRULENCE FACTORS)
Host range L. monocytogenes can transiently colonise the gut of humans and a wide range of animals, including cattle, sheep, goats, pigs, dogs, cats, domestic and wild rabbits, many other small mammals, and birds, including poultry.Clinical manifestations in animals include encephalitis, abortion, and sepsis [9].

Host response
The innate immune response occurs within minutes of the bacteria entering the body, activating bacteria encasement and lymphocyte, limiting systemic spread, and reducing the risk of life-threatening complications.The acquired immunity response involving specific CD4+ and CD8+ T cells mediates complete clearance of infection [8].Autophagy limits intracellular replication and pyroptosis of the infected cells limits the spread.Antibodies generated against L. monocytogenes can prevent reinfection.

Transmission
Transmission of L. monocytogenes through the consumption of contaminated foods (such as deli meats, cold cuts, hot dogs, fermented or dry sausages, unpasteurized milk and milk products, refrigerated pate, meat spreads, smoked seafood and unwashed fruits and vegetables), and more rarely through contact with infected animals [9].Vertical (mother-to-fetus) transmission is possible via the placenta during pregnancy and post-birth [2].

Infection
Ingested L. monocytogenes infects survives in the human gut and penetrates gut mucosa into the bloodstream to spread systemically.Invasive listeriosis is associated with severe disease outcomes, including bacteraemia, and respiratory infections in elderly immunocompromised patients.Listeria can cross the blood-brain barrier leading to meningitis, encephalitis, and neurolisteriosis.Listeria can also cross the placental barrier with long-term effects on the unborn [2].